Basic Science and Pathogenesis.

Background: Disrupted balance between amyloidogenic and non-amyloidogenic pathways leads to cognitive decline in Alzheimer's disease (AD). Evidence suggests vitamin A (VA) supplementation favors the non-amyloidogenic pathway through upregulation of α-secretase. Originally used to map embryonic retinoic acid (RA) signaling, RARE-LacZ mice possess multiple LacZ genes controlled by retinoic acid response elements (RAREs). We crossed RARE-LacZ mice with AD mouse models to determine their suitability for quantitative studies into the effects of VA on dentate gyrus (DG) RA signaling and learning in AD.

Methods: Relative LacZ gene copy ratio was determined by qPCR of LacZ in gDNA, normalized to ultra-conserved region 329. Dietary intervention compared VA supplemented (20 IU/g) AIN-93M to standard AIN-93M (4 IU/g VA). Mice were tested at postnatal day (P)125 via water T-maze (WTM, 9 simple discrimination, 9 reversal trials). PFA-fixed sections (40µm) were immunostained for LacZ, doublecortin, and/or calbindin, confocally imaged, and analyzed using ImageJ.

Results: RARE-lacZ mice were first crossed with C57BL/6J, -NJ, and CD1 mice (wildtype strains of J20, hAβ-KI, and RARE-LacZ mice, respectively). The relative LacZ gene copy ratio was ∼2.6:1 between RARE-LacZ mice (N = 12) and crosses (N = 34). Values for 32/34 offspring fell within ±50% of the mean. Strain affected latency to platform on WTM during simple discrimination and reversal (N = 11-12, p<0.05, Friedman), however total distance traveled was unaffected suggesting intact learning in all backgrounds. In WT strains, hippocampal LacZ immunoreactivity was localized to a subset of mature doublecortin-negative, calbindin-positive DG granule cells, appearing higher on C57BL/6J and -NJ than CD1 backgrounds. Offspring from J20 AD and RARE-LacZ mice exhibited impaired learning (N = 16, p<0.05, Kolmogorov-Smirnov). No significant difference in behavior between supplemented (20 IU/g) and standard (4 IU/g) VA was observed in AD mice. However, DG LacZ signal was completely silenced in 7/32 mice evaluated by immunostaining.

Conclusions: RARE-LacZ mice appear to have behavioral and genetic characteristics appropriate for testing VA-mediated interventions in AD models. RA signaling is prominent in mature DG cells associated with successful reversal learning. Although reversal of AD-related learning deficits by VA supplementation was not observed, testing AD mice on a VA deficient (0.4 IU/g) diet is planned.